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Determination of Protein Content in Soybeans Using the Kjeldahl Method
The Kjeldahl method, developed by Johan Kjeldahl at the Carlsberg Laboratory in Copenhagen, was originally designed for the brewing industry. However, its applications have since expanded significantly and are now widely used to determine nitrogen levels in various organic substances. These include foods, beverages, meat, agricultural products, animal feed, environmental water and soil samples, biological materials, dairy products, and pharmaceuticals.
Despite advancements and automation in the Kjeldahl method for total nitrogen determination, its fundamental principles have remained unchanged. The method comprises three primary steps: digestion, distillation, and titration. The process begins by converting the organic nitrogen in a sample into measurable ammonium ions, followed by titration analysis to determine protein content. Protein content is expressed as a percentage, calculated by multiplying the nitrogen result by a conversion factor derived from the average nitrogen content of proteins.
This article focuses on the determination of protein content in soybeans using BEING’s BPK-9870A Automatic Kjeldahl Nitrogen Analyzer, showcasing its precision and efficiency in modern protein analysis.
Soybeans and Their Nutritional Significance
Soybeans (Glycine max (L.) Merr.), a leguminous crop originally cultivated in China, were introduced to India for commercial production in the late 1960s. Within a decade, soybeans became a significant contributor to India’s oilseed production. They are consumed in various forms, including fermented foods (miso, tempeh, natto, soy sauce) and unfermented foods (tofu, soymilk, edamame, soy nuts, and sprouts).
Soybeans are renowned for their high nutritional value, serving as an excellent source of plant-based protein and oil. On average, they contain 40–50% protein and 18% oil, along with bioactive compounds like isoflavones, phytic acid, and saponins. Soybean proteins are classified based on their solubility into albumins (water-soluble), globulins (salt-soluble), glutelins (dilute acid/alkali-soluble), and prolamins (alcohol-soluble).
Efficient and accurate evaluation of protein content in food products is essential for assessing their nutritional and economic value.
BEING’s BPK-9870A Automatic Kjeldahl Nitrogen Analyzer
Experimental Procedure
1. Sampling
Nine varieties of soybeans were collected, and analyses were performed in triplicate to ensure accuracy.
2. Methodology
Protein content determination was carried out using the BEING BPK-9870A Automatic Kjeldahl Nitrogen Analyzer. The chemicals used were sourced from Sigma Aldrich and required no further purification. Approximately 0.5 g of each soybean variety was weighed and placed into 250 mL digestion tubes. Each tube received 20 mL of concentrated sulfuric acid and two catalyst tablets, each containing 5 g of potassium sulfate (K₂SO₄) and 0.5 g of copper sulfate pentahydrate (CuSO₄·5H₂O).
The digestion process was conducted at 400°C for three hours using a BRJX-8B Digestion System. Following digestion, samples were neutralized in a BZH-1B Acid-Alkali Neutralization Unit. Sodium hydroxide (32% NaOH) was added to the neutralized samples, and the resulting ammonia was distilled using the BPK-9870A Automatic Kjeldahl Nitrogen Analyzer. The distillate was collected in a known volume of standardized 0.1 N sulfuric acid (H₂SO₄).
Nitrogen content was then determined through titration with a standardized 0.1 N hydrochloric acid (HCl) solution, using a mixture of bromocresol green and methyl red as indicators. The crude protein content was calculated by multiplying the nitrogen content by the universally accepted conversion factor of 6.25. This factor assumes that proteins contain 16% nitrogen (100/16 = 6.25) and that all organic nitrogen originates from proteins.
The protein content of the nine soybean seed varieties analyzed is presented in Table 1.
| Sample Variety | Protein % |
|---|---|
| JS-335 | 40 |
| JS71-05 | 39 |
| PK - 564 | 41 |
| JS90 - 41 | 41 |
| JS75 - 46 | 38 |
| NRC - 2 | 39 |
| MACS58 | 39 |
| Monetta | 39 |
| MACS 13 | 38 |
Table 1: Percentage of protein content in sample soybean seeds
Conclusion
In an era where the purity and nutritional value of food are of growing concern, accurate protein analysis is crucial for both nutritional and economic assessments of food products. The Kjeldahl method remains an indispensable tool for nitrogen and protein determination due to its high accuracy, versatility, reproducibility, and universal acceptance.
The use of BEING’s BPK-9870A Automatic Kjeldahl Nitrogen Analyzer demonstrated efficient, accurate, and reproducible results in protein analysis for soybeans, reinforcing the method’s reliability and relevance in modern scientific research and industry applications.
References
1. https://iisrindore.icar.gov.in/statistics.html
2. Qin, P.; Wang, T.; Luo, Y. A Review on Plant-Based Proteins from Soybean: Health Benefits and Soy Product Development. Journal of Agriculture and Food Research 2022, 7 (7), 100265. DOI: 10.1016/j.jafr.2021.100265.
3. Friedman, M.; Brandon, D. L. Nutritional and Health Benefits of Soy Proteins. Journal of Agricultural and Food Chemistry 2001, 49 (3), 1069–1086. DOI:10.1021/jf0009246.